Effect Of Lactobacillus Reuteri On Intestinal Microbiota And Gene Expression Of Jejunum Tissues In Weaned Piglets

This study was performed to evaluate the biological characteristics of Lactobacillus reiteri ZLR003 in vitro.The effect of this strain on intestinal microbiota and jejunum transcriptome in weaned piglets were also determined.The study will reveal the mechanism of probiotic actions of Lactobacillus reuteri strain and promote its efficient utilization in pig production.In Exp.l,one Lactobacillus strain was isolated from cecum mucosa of healthy weaned piglet and screened with growth,acid tolerance,bile salt resistance and antibacterial properties.The strain was identified through standard morphological,biochemical and physiological tests,as well as by 16S rRNA gene sequence analysis and named as Lactobacillus reiteri ZLR003.L.reiteri ZLR003 strain was cultured in MRS medium with 18 h can be used as the best time for inoculation.The biological characteristics in vitro showed that L.reiteri ZLR003 strain could produce lactic acid and had stronger inhibition effect of E.coli.The strain had the ability of anti-oxidant.The viable counts of the strain at 70℃for 60 s could increase by 2.05 times when the strain was treated with 65℃ for 3 min.Glucose was the best carbon source of culture medium,and L-tryptophan could promte the growth of L.reiteri ZLR003 strain.In Expt.2,the effects of dietary supplementation with L.reuteri ZLR003 on the microbiota in weaned piglets of the V3-V4 hypervariable region of the 16S rRNA gene were investigated using Illumina Miseq technology.The results showed that treatment with L.reuteri ZLR003 increased the unique OTU numbers in jejunum compared with the chlortetracycline group.Proteobacteria was increased in the L.reuteri ZLR003,which was closest to that in the chlortetracycline group,while the L.reuteri ZLR003 group was difference from the chlortetracycline treatment at genus of microbiota.In colon and cecum,Treponema was significantly less abundant in the L.reuteri ZLR003 group than in the chlortetracycline and control groups.Distances between clustered samples revealed that the L.reuteri group was closer to the chlortetracycline group than the control group for jejunum samples,while colon and cecum samples of the L.reuteri ZLR003 group were clustered with those of the control group.In Expt.3,the gene expression in the jejunum tissue of weaned piglets was investigated by RNA-seq analysis after administering Lactobacillus reuteri ZLR003.In total,401 and 326 genes were significantly regulated in L.reuteri ZLR003 vs.control and L.reuteri ZLR003 vs.chlortetracycline,respectively.1083 and 900 GO terms and 78 KEGG pathways were measured in L.reuteri ZLR003 vs.control and L.reuteri ZLR003 vs.chlortetracycline,respectively.The GO terms related with"antioxidant ability" was significantly enrichment by L.reuteri ZLR003 when compared with the chlortetracycline and control groups.In addition,the KEGG pathway related to "arachidonic acid metabolism" and "linoleic acid metabolism" were enrichment after treatment with L.reuteri ZLR003 when compared with the other two groups.The fatty acid composition in the colon examined by Gas Chromatographic analysis and suggested that the MUFA C18:ln9c,and PUFAs C18:2n6c and C20:4n6 in colon were increased in the L.reuteri ZLR003 group,verifying the KEGG pathway analyses of the RNA-seq results.In conclusion,the strain L.reuteri ZLR003 exhibited excellent probiotic characteristics in vitro,including the ability to produce lactic acid,the stronger inhibition effect of E.coli and anti-oxidant.When weaned piglets treated with treatment with L.reuteri ZLR003,the unique OTU numbers were increased and the microbiota composition was affected at phylum and genus.At the level of gene expression,the results of RNA-seq showed that antioxidant ability and unsaturated fatty acid metabo/lism were improved by L.reuteri ZLR003.The results of this study would contribute to promote the application of the strain in pig production.